Activating mutations of the calcium-sensing receptor: The calcilytics ATF-936 and AXT-914 attenuate mutants causing autosomal dominant hypocalcemia and Bartter syndrome type 5

Introduction: Activating mutations of the calcium sensing receptor (CaSR) cause autosomal dominant hypocalcemia (ADH) characterized by low serum calcium, inappropriately low PTH and relative hypercalciuria. Four activating CaSR mutations cause additional renal wasting of sodium, potassium and other salts, a condition called Bartter syndrome type 5. Until today there is no medical treatment for Bartter Syndrome and ADH. We therefore investigated the effects of two novel calcilytics on activating CaSR mutants. Methods: 4 ADH mutations and all 4 known mutations causing Bartter syndrome type 5 were expressed in human embryonic kidney 293T cells and receptor signalling was studied by measurement of intracellular free calcium ([Ca 2+ ] i ) in response to different concentrations of extracellular calcium ([Ca 2+ ] o ). To investigate the effect of calcilytics, cells were stimulated with 3 mM [Ca 2+ ] o either in the presence or absence of NPS-2143 (300nM or 1µM), ATF-936 or AXT-914 (300nM each). Results: All eight mutants studied showed an elevated response to [Ca 2+ ] o with left shifted dose response curves as shown before. In contrast to NPS-2143, which was only partially effective, the novel calcilytics ATF-936 and AXT-914 significantly reduced the increased activity of all known Bartter syndrome type 5 and all studied ADH mutants. When these mutants were co-expressed with the wildtype receptor to approximate the situation in tissues of affected patients, ATF-936 and AXT-914 were equally effective on all mutants. Conclusion: Allosteric calcilytics like ATF-936 and AXT-914 are capable of attenuating the enhanced activity of CaSR mutations causing Bartter Syndrome type 5 and ADH. These new calcilytics might therefore offer a medical treatment option for patients with all types of activating mutations in the CaSR gene.